Medullary thymic epithelium (TE) participates in thymocyte negative selection. Heterogeneity of this TE compartment is centrally related to the ability of the medullary TE compartment to effect tolerance. The basis for this medullary TE heterogeneity is poorly understood, although the prevailing model holds that medullary TE exhibit "promiscuous" expression of tissue-specific genes as a consequence of a random and reversible derepression of gene expression that is regulated by the aire transcription factor. Studies proposed here will test the novel hypothesis that the heterogeneity of thymic epithelium reflects alternative developmental fates of progenitor epithelial cells, such that subsets of medullary TE differentiate to express molecules characteristic of other epithelial tissues. This model accounts for the patterns of medullary TE heterogeneity that have been previously observed and will allow an assessment of this heterogeneity in an organized and rational manner. Work proposed here will test the corollary hypotheses that tissue specific gene expression in the thymus is regulated in the same manner as in peripheral epithelial tissues. The work proposed here will also provide an understanding of how the medullary TE compartment develops and will provide new insight into the nature of the cellular interactions and signaling pathways that regulate the development of the these cells. Finally, studies proposed here will re-evaluate the impact of the aire transcription factor on the development of the organization and development of the medullary compartment. The questions addressed here are centrally related to a major question of clinical importance how the epithelial compartment contributes to the establishment of immunological tolerance to self-epithelial molecules. Understanding the mechanisms controlling the expression of potentially toleragenic antigens within the thymus will be necessary in order to design rationale therapeutic strategies to enhance this important aspect of thymic function. In addition to providing insight into this problem, validation of the model of medullary TE development proposed here will fundamentally alter the prevailing binary developmental model of TE development and provide new opportunities to enhance other aspects of thymic function that decline during age-related thymic involution or as a consequence of HIV infection.